Spatial Approximations between Residues 6 and 12 in the Amino-terminal Region of Glucagon-like Peptide 1 and Its Receptor A REGION CRITICAL FOR BIOLOGICAL ACTIVITY
Spatial Approximations between Residues 6 and 12 in the Amino-terminal Region of Glucagon-like Peptide 1 and Its Receptor A REGION CRITICAL FOR BIOLOGICAL ACTIVITY
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DOI：
10.1074/jbc.m110.135749
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发表时间：
2010-08-06
影响因子：
4.8
通讯作者：
Dong, Maoqing
中科院分区：
文献类型：
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作者：
Chen, Quan;Pinon, Delia I.;Dong, Maoqing
了解胰升糖素样肽1(GLP1)受体天然配体结合和激活的分子基础可能有助于开发用于治疗2型糖尿病的激动剂药物。我们以前报道过GLP1及其受体上的羧基末端残基24和35之间的分子近似。在这项工作中，我们重点研究了GLP1的氨基末端区域，这是已知的对受体激活至关重要的区域。我们开发了两个高亲和力、全激动剂的GLP1探针，在30个残基(GLP1(7-36))的第6位和第12位有共价连接位点。这两个探针都与受体特异性结合，并共价标记单个不同的位点。标记受体的化学和蛋白酶裂解将其氨基末端结构域的膜旁区域确定为第12位探针的共价结合区域，而第6位探针标记的区域定位于第一个细胞外环。放射化学测序表明，与第一跨膜片段相邻的受体残基Tyr(145)是第12位探针标记的部位，而第一细胞外环内的受体残基Tyr(205)是第6位探针标记的部位。这些数据为天然配体结合和激活BG家族蛋白偶联受体的共同机制提供了支持。多肽氨基末端结构域与受体相互作用的这一区域可能提供一个小分子激动剂靶向的口袋。
Understanding the molecular basis of natural ligand binding and activation of the glucagon-like peptide 1 (GLP1) receptor may facilitate the development of agonist drugs useful for the management of type 2 diabetes mellitus. We previously reported molecular approximations between carboxyl-terminal residues 24 and 35 within GLP1 and its receptor. In this work, we have focused on the amino-terminal region of GLP1, known to be critical for receptor activation. We developed two high-affinity, full agonist photolabile GLP1 probes having sites of covalent attachment in positions 6 and 12 of the 30-residue peptide (GLP1(7-36)). Both probes bound to the receptor specifically and covalently labeled single distinct sites. Chemical and protease cleavage of the labeled receptor identified the juxtamembrane region of its amino-terminal domain as the region of covalent attachment of the position 12 probe, whereas the region of labeling by the position 6 probe was localized to the first extracellular loop. Radiochemical sequencing identified receptor residue Tyr(145), adjacent to the first transmembrane segment, as the site of labeling by the position 12 probe, and receptor residue Tyr(205), within the first extracellular loop, as the site of labeling by the position 6 probe. These data provide support for a common mechanism for natural ligand binding and activation of family B G protein-coupled receptors. This region of interaction of peptide amino-terminal domains with the receptor may provide a pocket that can be targeted by small molecule agonists.